I completed my PhD in three years in the Department of Virology and Immunology of the Institute for Animal Health, England, where I studied the intracellular transport of African swine fever virus, a large DNA virus. My work led to the publication of 3 first-author papers. I graduated in September 2004. In January 2005, I joined Paul Bieniasz group at the Aaron Diamond AIDS Research Center, in New York, USA. As a post-doctoral fellow, I am continuing to explore the cell biology of virus infection. My current work focuses on the assembly and budding of Human immunodeficiency virus type I (HIV-1). I first established that the productive site of HIV-1 assembly was the plasma membrane. More recently, in collaboration with Sandy Simon, the head of the Cellular Biophysics laboratory at the Rockefeller University, we have developed innovative microscopic approaches to study HIV-1 particles assembly in real time, at the scale of individual particles. These approaches have allowed an unprecedented view of the genesis of individual virus particles in live cells and the determination of parameters that were inaccessible with conventional techniques. So far, my post-doctoral work led to the publication of 5 papers, including 3 first author papers. My immediate objectives are to exploit these approaches to understand the dynamic of the interaction between HIV-1 Gag proteins and key cellular factors at the site of particle assembly and release. During the K99 phase, I will spend more time in Sandy Simon laboratory, where I will be trained to develop new elaborate microscopic tools. I will pursue my work on the dynamic of HIV-1 assembly and release, using existing tools, as well as tools under development. Once the training phase of the grant will be completed, I intend to start my own group. As an independent investigator, I will continue exploiting the techniques and tools that I have developed during my post-doctoral training to pursue my studies on the cell biology of HIV-1 infection. Initially, during the R00 phase, I will focus on a particular aspect of HIV-1 replication: the cell biology of genome packaging. New approaches are definitively warranted in this field to determine, for instance, where in the cell the HIV-1 genome dimerizes, what mechanisms govern the cytoplasmic trafficking of the genome to assembly sites, where in the cell RNA first binds Gag, etc. This is also an area of research that is not studied in Paul Bieniasz laboratory, so my work as an independent investigator should not overlap with his. The R00 phase will allow me to start my own group without the burden of writing a grant in the first year and allow me to hire some staff immediately. After this grant-writing free time, my staff and I should have collected enough preliminary data to submit an R01. This first R01 should be a stepping-stone toward establishing a successful laboratory studying the cell biology of HIV-1. My long-term goal is to expand into other poorly understood areas of HIV-1 biology that interest me, such as on post-entry events. More specifically, I will focus my work on the formation, intracellular transport and nuclear import of partially disassembled viral particles, named pre- integration complexes. All cell biological aspects of these crucial steps of the HIV-1 life cycle are unknown;understanding them is likely to reveal new and fundamental cellular pathways. I am determined and highly motivated to pursue my career in the exciting field of HIV-1 cell biology. Receiving the competitive K99/R00 grant will certainly help me reach this goal. Project narrative: A better understanding of HIV-1 infection is crucial for the development of new classes of antiviral drugs. In order to dissect the mechanisms of HIV-1 assembly, we have developed an assay that allows us to visualize the formation of viral particles in real time and at the scale of single particles. We propose to use this innovative assay to study the dynamic interactions between key viral and cellular components during HIV-1 assembly and release.